Embryonic limbs developing in organ culture have been shown to be capable of the same pathological responses to a number of teratogens as limbs developing "in vivo." Application of this procedure for teratogenic screening is limited by the absence of cytochrome P-450-metabolizing activity in the culture environment. We propose to add hepatic microsomal enzymes to limb bud culture to effect metabolism of compounds requiring P-450 activation. Our first objective is to utilize cyclophosphamide as a model compound to evolve procedures necessary to achieve cytochrome P-450 metabolism "in vitro" and delivery of metabolites to the developing limb. These procedures will be used to elucidate the mechanism of cyclophosphamide-induced teratogenesis, as well as the conditions related to maternal and fetal metabolism of cyclophosphamide. Our second objective is to apply the model system to environmentally-related compounds such as benzo(a)pyrene, diethylstylbesterol, and tap water concentrates to investigate the influence of P-450 metabolism on their teratogenic activity.